More on B12 content in Tempeh, Seaweeds, Organic Produce, and Other Plant Foods, below…

Summary: No food in Europe or the U.S. other than Japanese Nori, has been tested* for lowering MMA levels (*at the time of publication of this original article).

Thus, the discussion about whether Western vegans can get B12 from plant foods can, and probably should, end here (until proper research is conducted). Because so many plant foods have failed other tests that do not measure up to the MMA lowering test, and because there are so many false rumors being passed around, the studies of B12 in plant foods are examined in detail below.

The only plant foods which have been tested for B12 activity using the gold standard of lowering MMA levels in humans are dried and raw nori from Japan. Dried nori made MMA status worse, indicating that it can reduce B12 status and can possibly harm people who are B12-deficient. Raw nori kept MMA levels about the same, indicating that it didn’t harm B12 status, but it did not help either.

Of all the foods studied below, only tempeh in Indonesia or Thailand, dulse, Chlorella, raw nori, Aphanizomenon flos-aquae, and coccolithophorid algae warrant much further attention for providing B12. Unless these foods are shown consistently to correct B12 deficiency, vegans should not rely on them as a B12 source.

It would be great to find a reliable plant source of B12 for vegans. One might get the opposite impression given my level of critique of some of these studies. My skepticism is only due to the potential harm from relying on a food that contains inactive B12 analogues which can make a B12 deficiency even worse.

There has been a long history of misconceptions about which, if any, plant foods are sources of B12. Much of this stems from the methods of measuring B12 analogues. Other confusion stems from bacterial contamination that occurs in some foods but not others. Please see Measuring B12 in Plant Foods: Why the Confusion? for an explanation of the methods for for measuring B12 analogues in plant foods.

Unlike animals, most, if not all, plants have no B12 requirement for any function, and therefore have no active mechanisms to produce or store B12. Many seaweeds have been shown to have B12 analogues.

Most seaweeds are macroalgae, which are technically not plants. Some microalgae contain an enzyme that can use cobalamin, but also have an enzyme with the same function that does not require cobalamin in case it is not present. These macroalgae do not make their own cobalamin, but rather have a symbiotic relationship with cobalamin-producing bacteria (1). Note that I am purposefully using the term “cobalamin” rather than “vitamin B12” because it is not clear if these cobalamins are active vitamin B12 in humans.

During the 1970s, two enzymes in plants (potatoes and bean seedlings) were found to respond to the addition of adenosylcobalamin (2, 3) (a co-enzyme form of B12). One explanation is that adenosylcobalamin provides some factor that is usable by these enzymes, but that adenosylcobalamin is not required by these plants for growth. Thus far, these plants have not been shown to counteract B12 deficiency symptoms (though I am not aware of any well-designed attempts as it is assumed that they do not contain B12). It is probably safe to assume that many vegans who have developed severe B12 deficiency ate potatoes and beans.

There are some rumors, though no evidence of which I am aware, that if you let organic produce, such as carrots, sit at room temperature for a few hours, bacteria on the surface of the carrots will produce B12. For this to happen, specific species of bacteria would be required, as would cobalt. Until there is research showing that such a method can lower MMA levels, such produce should not be considered to provide B12.

Various studies have tested the foods in Table 1 for B12 analogues and found none. To my knowledge, other than in Mozafar’s studies (below) in which B12 and cow dung were carefully added to the soil of potted plants, no published study has shown any B12 analogues in any of these foods.

As you can see, there are very small amounts, if any. Since the amounts are so small, any inactive analogues should not significantly interfere with an individual’s active B12 from other sources, and if the analogue is active B12, it will not provide much. Thus, these foods should neither add to, nor detract from, a vegan’s B12 status.

Tempeh

For a long time, tempeh has been said to contain B12. Table 3 shows the results of measuring B12 analogue in various tempehs.

Table 3. B12 Analogue Content (µg/30 g) of Tempehs

Netherlands4

USA7

Indonesia5,6

Assay

IF

IF

IF or R-proteinA

tempeh

ND

.02C

.054-1.2B

A – Used an assay method by Lau et al.32 (1965) which uses R-protein or IFB – 10 commercial tempeh samples purchased from various markets in Jakarta, Indonesia

C – Cooked for 60 minutes

IF – Intrinsic factor

ND – None Detected

The studies in the USA and in The Netherlands showed little to no B12 analogue.

In contrast, Areekul et al. (6) (1990, Indonesia/Thailand) found more significant amounts of B12 analogue. Tempeh production requires molds belonging to the genus Rhizopus. These were found not to produce B12 analogues in Areekul et al.’s study. Rather, a bacterium, identified as Klebsiella pneumoniae, was isolated from the commercial tempeh starter and determined to be the B12 analogue source. This confirmed Albert et al.’s (8) (1980) finding that the Klebsiella genera could produce B12 analogues. In Albert’s study, the analogue was thought to be active B12. Whether the analogues found by Areekul et al. were the same as in Albert’s study is not known. Given that K. pneumoniae is not required for tempeh production, we can conclude that the B12 analogue found in the tempehs in Indonesia were due to bacterial contamination (though apparently common there). Tempeh in Europe and the U.S. cannot be relied on as a source of B12. Until tempeh in Indonesia is shown to reduce MMA levels, it should not be relied upon there, either.

Blue-Green Algae

Blue-green algae are also known as cyanobacteria, blue-green bacteria, and cyanophyta. They are not actually algae, but rather organisms with characteristics of both bacteria and algae. They can peform photosynthesis and are thought to be the ancestors to chloroplasts in algae and plants.

“Is the vitamin B12 in SBGA bioavailable and bioactive? Yes. The Super Blue Green Algae (SBGA) strain, Aphanizomenon flos-aquae, has been tested by Lancaster Labs for B12 analog levels using microbiological testing methods that are comparable to methods 952.20 and 960.46 of the Association of Analytical Chemists (AOAC). Unlike other plant foods such as Spirulina, which contain corrinoids with virtually no vitamin B12 activity, Aphanizomenon flos-aquae is a reliable source for vegetarians seeking to supplement their diets with a bioactive form of this important nutrient.”

However, test methods 952.20 and 960.46 use Lactobacillus leichmannii (9), which can measure non-B12 corrinoids (10). See the table Test Organisms for B12 Microbiological Assays in Measuring B12: Why the Confusion? Thus, it can only be concluded that Cell Tech’s SBGA contains B12 analogues whose activity is yet to be determined.

2010 Update: It appears that Cell Tech is now the company, Simplexity Health, and is no longer touting SBGA as a source of vitamin B12.

In a 2009 study from Italy (11), researchers gave Aphanizomenon flos-aquae to 15 vegans. First there was a washout period in which the vegans took no supplemental B12 for 3 months. They were then given 6 capsules of Klamanth Algae from Nutratec (which also contained digestive enzymes to help absorption).

The results, seen in Table 4, show that the average homocysteine level went down. The authors believe this is an indication that Aphanizomenon flos-aquae is a source of active vitamin B12, and that it “warrants further larger, and longer-term randomized trials to confirm such preliminary conclusions.”

Here are some problems with the study:

The authors state in the paper that homocysteine is the most reliable marker for B12 activity, but it is not. Homocysteine levels can be affected by folate intake and, to a lesser extent, vitamin B6. Methylmalonic acid levels are the most reliable marker for B12 activity. This is well known and uncontroversial, so it is odd that the researchers did not know this.

The authors noted that vitamin B6 could not have reduced the homocysteine levels because the algae has very little. They also said that folate levels could not have affected them, but in looking at the results, folate levels did increase (even though the difference was not statistically signifcant).

The homocysteine levels of these vegans started out pretty high, and when the study ended they were still much too high. A safer level is closer to 6 – 8 µmol/l.

One subject’s homocysteine level increased, and one subject’s homocysteine level that was about 10 µmol/l did not respond to the aglae supplementation.

The researchers obtained the algae directly from a company that produces it. It would have been more reassuring if the algae were purchased in a store where the company didn’t realize it was going to be tested.

In another study from Italy (2002) (12), vegetarians had really high homocysteine levels (25 µmol/l). This is much higher than almost all other studies, which makes one wonder what’s going on in Italy.

In conclusion, it appears that Aphanizomenon flos-aquae might provide some vitamin B12 activity in humans. On the other hand, it did not succeed in lowering homocysteine to an ideal level whereas vitamin B12 supplements do succeed at doing so. At this time, it would be prudent not to rely on it for optimal health.

The wide range of B12 analogues from one measurement method to another indicates that spirulina has a wide variety of different analogues, many of which are inactive. Some may interfere with B12 activity in humans.

Table 6 shows the B12 analogue content of arame, dulse, hijiki, kelp, kombu, and wakame per 30 g of seaweed. Please note that 30 g is a lot of seaweed. A serving size would be closer to 3 grams. Seaweeds also tend to be very high in iodine, which can cause problems at high intakes. So, consuming mass quantities of seaweed is unadvisable.

The only seaweed in this list that warrants further study is dulse (also spelled “dulce”), which contains .3 to .39 µg of B12 analogue per 3 g serving. Unless dulse is eventually shown to lower MMA levels, it should not be considered a source of active B12.

Chlorella

Pratt & Johnson (15) (1968, USA) studied numerous batches of chlorella and occasionally found amounts of B12 analogue that were in the range of error for the test method. In other words, they were not able to detect practical amounts. They noted that their extraction processes might not have been adequate though they used many different methods. They also noted that their synthetic medium on which the chlorella was grown might have interfered with B12 analogue synthesis.

Kittaka-Katsura et al. (16) (2002, Japan) measured B12 analogue levels in Chlorella using both a Lactobacillus leichmannii ATCC 7830 and an intrinsic factor assay. Both methods showed about the same amount of B12 analogue, listed in Table 7 below:

Chen and Jiang (17) (2008, Taiwan) used capillary electrophoresis to detect cyanocobalamin and hydroxocobalamin in chlorella. Capillary electrophoresis is a relatively new method that should be able to detect the exact structure of a cobalamin analogue. They found considerable amounts of cyanocobalamin in two samples of Chlorella, with negligible amounts of B12 analogues.

Until chlorella is tested on humans to determine whether it lowers MMA levels, it should not be considered a reliable source of vitamin B12, especially since the study by Pratt & Johnson (15) showed no vitamin B12.

Various batches of nori were found to contain significant amounts of B12 analogue. One study verified the molecular weight through paper chromotagraphy, indicating that there is a good chance that some of this B12 is active. Yamada et al. (20) (1996, Japan) determined that nori contains what they considered to be active B12 analogues using various assays and methods (results not reported here). However, in light of Dagnelie’s results, Yamada et al. (19)(1999, Japan), decided to test the nori to see if it could reduce methylmalonic acid (MMA) levels, the gold standard for determining the B12 activity of a food:

Raw nori was purchased within 48 hours of harvesting. Dried nori was purchased from a store. Inactive vs. active B12 was determined by IF assay and confirmed by paper chromatography. 10 people (all nonvegetarian) were then studied. The results are shown in Table 9.

The results indicate that B12 in raw nori can be changed into harmful inactive B12 analogues by drying, and that dried nori decreases B12 status. Yamada et al. said that although dried nori cannot be used as a B12 source, in small amounts it is not harmful. However, they believe that raw nori is an excellent source of genuine B12.

I disagree with their conclusion that raw nori is an excellent source of active B12. While eating raw nori, the subjects’ uMMA levels increased 5%. While this was not enough of an increase to be statistically significant, it indicates that the raw nori did not improve B12 status (which would have required MMA levels to drop, rather than increase). This study showed that this batch of raw nori did not have enough inactive B12 versus active B12 analogue to be considerably detrimental, but it did not prove any benefit.

The study by Yamada et al. was further confounded by adding valine (an amino acid that can be converted into MMA when B12 is deficient) to the subjects’ diet in order to increase MMA levels so that a difference could be seen. The valine did not appear to do this when given without the nori, and no control groups were included, making the results even more difficult to interpret.

Other studies have measured the B12 analogue content of nori, but without testing to see if they could lower MMA levels:

A – Study said the amount was “identical” to that found with IF; the number was not actually givenA – Equivalent amounts

IF – Intrinsic Factor

Coccolithophorid algae (Pleurochrysis carterae) is being used in Japan as a calcium supplement. Miyamoto et al. (21) (2001, Japan) analyzed it for B12 analogue content:

They tried to verify whether it is an active form of B12 through liquid chromatography, and thought that is was. B12 analogue remained stable for 6 months of storage. They later followed up with a second study on coccolithophorid algae (22), but still did not test it to see if it can lower MMA levels in humans.

This algae deserves further attention to see if it can lower MMA levels.

A Case of False Reporting on the Benefit of Seaweed and Fermented Foods

Specker et al. (7) (1988, USA) reported a macrobiotic mother of an infant with a uMMA of 146 µg/mg who modified her diet by increasing her consumption of seaweeds and fermented foods. The infant’s uMMA dropped to 27 µg/mg in 2 months and to 13 µg/mg in 4 months. It was later discovered that this mother had also eaten fish and clam broth which were probably responsible for the improvement rather than the seaweeds and fermented foods (23). Specker et al. stated, “The vegetarian community we worked with believed fermented foods in their diet contained adequate amounts of vitamin B12.” However, on analysis, the fermented foods were shown not to have B12. (7).

Genmai-Saishoku Paradox?

Suzuki (24) (1995, Japan) studied 6 vegan children eating a genmai-saishoku (GS) diet, which is based on high intakes of brown rice and contains plenty of sea vegetables, including 2-4 g of nori per day (“dried laver”); as well as hijiki, wakame, and kombu. The foods are organically grown and many are high in cobalt (buckwheat, adzuki beans, kidney beans, shiitake, hijiki). Serum B12 levels of the children are shown in Table 11:

A – Exclusively breast-fed until 6 months old. Mothers had been vegan for 9.6 and 6.5 yrs prior to conception. Both mothers consumed 2 g of nori per day.

None of the many measurements between the vegans and 4 nonvegan controls were significantly different, including serum B12, MCV, and iron indicators. MMA and homocysteine levels were not measured. Some suggestions as to how the vegans got their B12 are:

From nori or the other seaweeds. The nori was most likely dried.

Small amounts of B12 from B12 uptake or contamination of plants grown in manure.

B12 from their mothers’ stores.

These results are both interesting and perplexing. The serum B12 levels are easy to explain as possibly being inactive B12 analogues. But it is particularly impressive that the eight-year-olds were doing well given that their mothers had been vegan for some time, supposedly without B12-fortified foods or supplements. Unfortunately, many vegan children have not had the same positive results, and until more is known about the GS children’s diets, this study should be considered an unsolved mystery.

If these children were my own, I would make sure they started to get at least a modest B12 supplement to ensure their continued good health.

Soil and Organic Produce as a B12 Source for Vegans

It is common in vegan circles to hear that if your produce has soil on it and you do not wash the produce before eating it, bacteria that lives in the soil and on the produce will provide B12. It is also claimed that in today’s world, our food supply is very sanitized whereas in the past, vegan humans would have received plenty of B12 from the unsanitized produce. What is the evidence for these claims?

B12 Analogue in Soil

There is a one paragraph report often cited in vegan literature for showing that B12 is found in the soil. Robbins et al. (25) (1950, New York) used Euglena gracilis var. bacillari as a microbiological assay for vitamin B12 “or its physiological equivalent.” A considerable proportion of bacteria and actinomycetes (molds) in the soil were found to synthesize B12 analogues. B12 analogues were also found in the roots of plants (.0002-.01 µg B12/g of fresh material). Some stems had some B12 analogue, but leaves and fruit generally did not. B12 analogue was also found in pond water and pond mud. There was no indication in the report as to how many different soils were tested, but the impression was that it was all in one local area. There is no way to know whether these molecules were active or inactive B12 analogues.

Iranian Villagers

Herbert (26) reported a group of “vegan” Iranians growing plants in night soil (human manure). The vegetables were eaten without being carefully washed and the amount of B12 was enough to prevent deficiency. However, for this information, Herbert cites Halstead et al. (1959) (27), who do not mention these Iranians in their paper. Herbert possibly meant to cite a 1960 paper by Halstead et al. (28) which reported that some Iranian villagers with very little animal product intake (dairy once a week, meat once a month) had normal B12 levels. None had megaloblastic anemia. Their average B12 level was 411 pg/ml which was quite high considering their diet. The authors speculated this could be because their diets, which were very low in protein, allowed for B12-producing bacteria to ascend into the ileum where the B12 could be absorbed. They also speculated that because they lived among their farm animals and their living areas were littered with feces, they picked up enough B12 through contamination.

Organic Produce as a B12 Source for Vegans

Soybean Plants Absorb B12

Mozafar & Oertli (29) (1992, Switzerland) added cyanocobalamin to the soil of soybean plants in amounts ranging from 10 to 3200 µmol/l. Using an intrinsic factor assay, 12-34% of the B12 was absorbed by the plants. 66-87% of the absorbed vitamin remained in the roots and the rest was transported to the various other parts, mainly the leaves. Mozafar points out that the concentrations of B12 in the soil used in this study were many times higher than the reported vitamin concentration in soil solution (.003 µmol/l) measured by Robbins (25).

In light of the above results, Mozafar (30) (1994, Switzerland) then studied how the B12 levels in plants are affected by adding cow dung to the soil. An assay using pig intrinsic factor was used to measure the B12 analogue. The study looked at the B12 analogue content of both organically fertilized soil and plants.

Two samples were taken from soil that had been treated with organic fertilizer every 5 years over the previous 16 years. The B12 analogue content in these samples was compared to soil that had only synthetic fertilizer applied. Results are shown in Table 12.

A,B – Statistically significant difference between groups with same letters

Soybean, barley, and spinach plants were then grown in pots of 2.5 kg of soil. 10 g dry cow manure was added to each pot. Plant parts were thoroughly washed to remove any soil before B12 was measured. Table 13 shows the results.

Further analysis showed that most or all of the B12 analogue in the plants was unbound. Mozafar concluded that plant uptake of B12 from the soil, especially from soil fertilized with manure, could provide some B12 for humans eating the plants, and may be why some vegans, who do not supplement with B12, do not develop B12 deficiency.

Does this mean that organic foods are a good source of B12? No. These studies show that when B12 analogues are placed in the soil, plants can absorb them.

Mushrooms and B12

Table 14. B12 in Mushrooms

Button

Cup

Flat

Most

Cap

1005

567

161

Flesh

233

83

84

Stalk

17

255

465

Peel

217

1015

354

Total (ng / 400 g)

1472

1920

1064

ng / Cupa

257.60

336.00

186.20

mcg / Cup

0.26

0.34

0.19

Cups to meet RDA

9.32

7.14

12.89

Least

Cap

11

8

17

Flesh

4

7

4

Stalk

11

7

12

Peel

36

20

68

Total (ng / 400 g)

62

42

101

ng / Cupa

10.85

7.35

17.68

mcg / Cup

0.01

0.01

0.02

Cups to meet RDA

221.20

326.53

135.79

aAssume 70 g per Cup

In 2009, a paper was published looking at the B12 analogue content of mushrooms in Australia (31). The authors used chromatography and mass spectrometry to determine whether the B12 was an active form, and they believed that it was.

Table 14 shows the B12 analogue content of the batches of each mushroom containing the most B12 and the batches containing the least.

Assuming that the B12 is active analogue, it would take anywhere from 7 to 326 cups of mushrooms to meet the RDA.

As for the source of the B12, the authors were not sure, but they said:

The high concentration of vitamin B12 in peel suggests that it was not synthesized within the mushrooms but was either absorbed directly from the compost or synthesized by bacteria on the mushroom surface. The latter is more likely because mushrooms have no root system to take up the vitamin in the compost as is the case with the uptake of vitamins by root plants from the soil containing fertilizers.

Conclusion About Organic Produce as a B12 Source for Vegans

Unless uncleaned, organic produce is shown to lower MMA levels, it is unjustified to claim that B12 can be obtained in such a manner, or to claim with certainty that humans have ever relied on it as a source of B12.

Only until organic foods are chosen randomly from markets and grocery stores throughout the country (or world) and are consistently shown to decrease MMA levels will someone not be taking a considerable risk in relying on organic foods for B12. This article documents many vegans suffering from B12 deficiency, and it is safe to assume that many of them consumed significant amounts of organic foods.

Additional note: The vegan movement is typically not aiming for a world where there are enough cows to produce a significant amount of manure for fertilizer.

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